ABSTRACT: The central nervous system (CNS) is home to three major classes of glia: astrocytes, microglia, and oligodendrocytes. In the last two decades, a fourth class of glia has been increasingly recognized, called oligodendrocyte progenitor cells (OPCs). Like microglia, OPCs have a stellate morphology and are ?tiled?, spanning defined territories across the gray and the white matter [7]. For a long time, our vision of OPC function was limited to the generation of mature oligodendrocytes, but this dogma is starting to be challenged by recently published studies. In the adult CNS, OPCs are the most proliferative cell type, a property not consistent with the need to generate new oligodendrocytes, since only a small percentage of OPCs is needed to give rise to oligodendrocytes in the adult, suggesting that OPCs could serve other role(s) in the CNS. Indeed, a nascent body of work is showing a more intricate role for these cells. For example, OPCs have been shown to receive GABAergic and glutamatergic synaptic input [8, 9], suggesting that they could be capable of sensing and modulating network activity. OPC depletion in the brain can trigger depressive-like behavior, placing this cell as an active participant in the etiology of depression [10]. Under pathological conditions, OPCs are actively recruited to injury sites, where they orchestrate glial scar formation and contribute to the immune response [11-13]. Taken together, these data demonstrate that OPCs are not simple bystanders but could participate in many CNS functions. Our goal with this proposal is to test the hypothesis that unique subsets of OPCs exist in the CNS and that they exhibit a distinct molecular signature. The foundation for this proposal is our preliminary data showing that more than one population of OPCs exists in the adult CNS, as shown with single cell sequencing. Guided by this evidence, our hypothesis will be addressed by pursuing two specific aims: 1) Explore OPC diversity in the adult CNS under homeostatic and pathological conditions. 2) Validate subsets of OPCs in vivo by analyzing expression of unique markers. Under the first aim, using our validated OPC reporter mouse strain as a tool to isolate OPCs, we will perform single cell sequencing analysis to discover the molecular diversity of OPCs. We have chosen to perform these studies using homeostatic as well as pathological conditions in which OPC activation occurs, indicated by their proliferation. In the second aim, we will validate our discovery of unique OPC subsets in the adult brain by performing staining for unique markers and begin to explore the function of these subsets. Our proposal is novel because we propose to discover new functions of OPCs using a state-of-the-art method (single cell sequencing). Our proposal is significant because these studies will provide new insight into the function of OPCs beyond remyelination.